Fig. 2

Nicotinamide increases total mitochondria within retinal ganglion cells axons in the optic nerve, which are not lost with complex I inhibition. (A) In MitoV mice, high resolution imaging of the optic nerve longitudinal sections allowed RGC specific YFP + mitochondria in RGC axons and somas to be captured for 3D reconstruction. (B) Morphological analysis revealed no significant changes in morphology induced by rotenone, with only a small increase in individual mitochondria volume in NAM treated, uninjured animals (DMSO + NAM). (C) Nicotinamide also significantly increased the density of RGC mitochondria in the optic nerve (number mitos/field) which remained high even with rotenone induced complex I inhibition. (D) To investigate mitochondrial changes at the level of individual RGC axons, we intravitreally injected an AAV2.2 encoding mCherry under a CMV promotor at low titer to sparsely label RGC axons. Individual mitochondria within these axons were reconstructed. (E) These mitochondria followed the same trend as for the optic nerve globally, but rotenone did induce a morphological change towards more spherical mitochondria, diverging from the typical prolate morphology of RGC axonal mitochondria. This did not occur in NAM treated animals, with uninjured NAM treated animals (DMSO + NAM) demonstrating that NAM promotes an exaggerated prolate morphology in the optic nerve. * P < 0.05, ** P < 0.01, *** P < 0.001, NS = P > 0.05